Structure-specific binding of the two tandem HMG boxes of HMG1 to four-way junction DNA is mediated by the A domain

Michelle Webb, Jean O. Thomas

    Research output: Contribution to journalArticlepeer-review

    Abstract

    We have investigated the nature of the 'structure-specific' binding of the tandem A and B HMG boxes of high mobility group protein 1 (HMG1) to four-way junction DNA. AB didomain binding favours the open, planar form of the junction, as shown by reaction with potassium permanganate. Site-directed cleavage of the DNA by a 1,10-phenanthroline-copper moiety attached to unique natural or engineered cysteine residues in the A or B domain shows that the two linked HMG boxes are not functionally equivalent in four-way junction binding. The A domain of the didomain binds to the centre of the junction, mediating structure-specific binding; the concave surface of the domain interacts with the widened minor groove at the centre, contacting one of the four strands of the junction, and the short arm comprising helices I and II and the connecting loop protrudes into the central hole. The B domain makes contacts along one of the arms, presumably stabilising the binding of the didomain through additional non-sequence-specific interactions. The isolated B domain can, however, bind to the centre of the junction. The preferential binding of the A domain of the AB didomain to the centre correlates with our previous finding of a higher preference of the isolated A domain than of the B domain for this structurally distinct DNA ligand. It is probably at least partly due to the higher positive surface potential in the DNA-binding region of the A domain (in particular to an array of positively charged side-chains suitably positioned to interact with the negatively charged phosphates surrounding the central hole of the junction) and partly to differences in residues corresponding to those that intercalate between bases in other HMG box/DNA complexes.
    Original languageEnglish
    Pages (from-to)373-387
    Number of pages14
    JournalJournal of molecular biology
    Volume294
    Issue number2
    DOIs
    Publication statusPublished - 26 Nov 1999

    Keywords

    • HMG box
    • N-(1,10-phenanthroline-5-yl)iodoacetamide
    • Potassium permanganate
    • Site-specific DNA cleavage

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